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Title: Improving peppermint essential oil yield and composition by metabolic engineering

Author
item LANGE, B.M. - Washington State University
item MAHMOUD, S.S. - Washington State University
item WILDUNG, M.R. - Washington State University
item TURNER, G.W. - Washington State University
item DAVIS, I.L. - Washington State University
item BAKER, R.C. - Washington State University
item Boydston, Rick
item CROTEAU, R.B. - Washington State University

Submitted to: Proceedings of the National Academy of Sciences (PNAS)
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/26/2011
Publication Date: 10/21/2011
Citation: Lange, B., Mahmoud, S., Wildung, M., Turner, G., Davis, I., Baker, R., Boydston, R.A., Croteau, R. 2011. Improving peppermint essential oil yield and composition by metabolic engineering. Proceedings of the National Academy of Sciences. 41:16944-16949.

Interpretive Summary: In the present study we have investigated the utility of overexpressing genes that encode enzymes involved in the pathway for monoterpene biosynthesis. We have generated transgenic plants, tested under greenhouse and commercial-scale field trial conditions, that express postive affects on both oil yield and composition. We have also produced transgenic lines containing a chemical marker, which enables the tracing of oil distilled from transgenic plants without any adverse effect on oil yield. Our data provide evidence that oil yield and composition in transgenic lines are stable for multiple growing seasons. Overall, our metabolic engineering approaches to modulate the expression of genes involved in essential oil production were highly successful. This research has yielded valuable insights that now enable the knowledge-based metabolic engineering of other commercially important essential oil plants.

Technical Abstract: Peppermint (Mentha x piperita L.) was transformed with various gene constructs to evaluate the utility of metabolic engineering for improving essential oil yield and composition. Oil yield increases were achieved by overexpressing genes involved in the supply of precursors through the 2C-methyl-D-erythritol 4-phosphate (MEP) pathway. Two-gene combinations to enhance both oil yield and composition in a single transgenic line were assessed as well. The most promising results were obtained by transforming plants expressing an antisense version of (+)-menthofuran synthase, which is critical for adjusting the levels of specific undesirable oil constituents, with a construct for the overexpression of the MEP pathway gene 1-deoxy-D-xylulose 5-phosphate reductoisomerase (up to 61% oil yield increase over wild-type controls with low levels of the undesirable side-product (+)-menthofuran and its intermediate (+)-pulegone). Elite transgenic lines were advanced to multi-year field trials, which demonstrated consistent oil yield increases of up to 78% over wild-type controls and desirable effects on oil composition under commercial growth conditions. The transgenic expression of a gene encoding (+)-limonene synthase was used to accumulate elevated levels of (+)-limonene, which allows oil derived from transgenic plants to be recognized during the processing of commercial formulations containing peppermint oil. Our study illustrates the utility of metabolic engineering for the sustainable agricultural production of high quality essential oils at a competitive cost.